DNA methylation is a type of chemical modification of DNA that can be inherited and subsequently removed without changing the DNA sequence. As such, it is part of the epigenetic code (Jaenisch & Bird. (2003) Nature Genetics, 33, 245, herein incorporated by reference in its entirety). DNA methylation involves the addition of a methyl group to a DNA nucleobase. In the most common example, a methyl group is added to the number 5 carbon of the cytosine pyrimidine ring. Cytosine methylation generally has the effect of reducing gene expression.
Methylation is a common capability of all viruses for self/non-self identification. DNA methylation at the 5 position of cytosine has been found in every vertebrate examined. In adult somatic tissues, DNA methylation typically occurs in a CpG dinucleotide context; non-CpG methylation is prevalent in embryonic stem cells (Dodge et al. (2002) Gene 289 (1-2): 41-48., Haines et al. (2001) Developmental Biology 240 (2): 585-598., herein incorporated by reference in their entireties). In plants, cytosines are methylated both symmetrically (CpG or CpNpG) and asymmetrically (CpNpNp). Long term memory storage in humans may be regulated by DNA methylation (Miller & Sweatt. (2007-Mar.-15) Neuron 53 (6): 857-869., Powell & Devin. (2008) New Scientist., herein incorporated by reference in their entireties).
In mammals, DNA methylation is essential for normal development and is associated with a number of key processes including imprinting, X-chromosome inactivation, suppression of repetitive elements and carcinogenesis. Between 60-90% of all CpGs are methylated in mammals (Tucker. (2001) Neuron. 30(3): 649-52., herein incorporated by reference in its entirety). CpGs are grouped in clusters called “CpG islands” that are present in the 5′ regulatory regions of many genes. In many disease processes such as cancer, gene promoter CpG islands acquire abnormal hypermethylation, which results in heritable transcriptional silencing.
Methods for improving sensitivity and robustness of methylation detection assays are needed.